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Implementing differentiated instruction (DI) into the classroom is a major challenge for beginning teachers. This study investigated which individual (self-efficacy, autonomy) and professional learning communities (PLCs) characteristics (collective responsibility, reflective dialogue, deprivatized practice) influence beginning teachers’ professional development in DI. From 65 primary schools, 227 beginning teachers participated. Path analyses showed that individual characteristics and reflective dialogue directly predicted teachers’ changes in DI-practice. Moreover, autonomy, collective responsibility, and reflective dialogue indirectly predicted changes in DI-practice via self-efficacy. Findings suggest that, to promote DI-implementation, beginning teachers could benefit from a joint sense of responsibility toward student learning and in-depth conversations with colleagues. Importantly, collegial support and autonomy could potentiate self-efficacy which may in turn stimulate the use of diverse instructional strategies.